Macrophage erythropoietin receptor signaling drives macrophage-myofibroblast transition via JAK2/STAT3 and TGF-β1/Smad3 pathways to exacerbate pulmonary fibrosis in mice.

The macrophage-myofibroblast transition (MMT) plays a significant role in the pathogenesis of pulmonary fibrosis; however, the regulatory mechanisms underlying this process are not fully elucidated. In this study, we provide evidence that signaling through the erythropoietin receptor (EPOR) in macrophages promotes MMT and exacerbates fibrotic progression. Through an integrated analysis of single-cell data from human idiopathic pulmonary fibrosis (IPF) samples (GSE136831), we identified the presence of MMT characterized by increased EPOR expression in macrophages. In experimental models of bleomycin-induced pulmonary fibrosis, mice with macrophage-specific EPOR knockout (cKO) exhibited enhanced survival, reduced MMT, and decreased collagen deposition compared to wild-type (WT) counterparts. Conversely, mice overexpressing EPOR (cKI) showed diminished survival, intensified MMT, and worsened fibrotic pathology. Mechanistically, EPOR activation in bone marrow-derived macrophages (BMDMs) stimulated with TGF-β1 enhanced TGF-β1/Smad3 and JAK2/STAT3 signaling pathways, while the JAK2 inhibitor AG490 inhibited EPOR-mediated Smad3 activation. BMDMs from EPOR-cKI mice displayed increased M2 polarization, and conditioned medium derived from MMT elevated the expression of α-SMA and collagen-1 in fibroblasts. Notably, the Smad3 agonist SRI-011381 reversed the suppression of MMT observed in EPOR-cKO macrophages. while the Smad3 inhibitor (SIS3) abolished EPOR-cKI-induced MMT responses. Our findings identify macrophage EPOR signaling as a regulator of MMT via the activation of JAK2/STAT3 and TGF-β1/Smad3 pathways, suggesting a potential novel therapeutic target for the treatment of fibrotic lung diseases.